Proteinase secretion in serum-independent cell lines established from human cancer cell lines.

1.非血清依赖性人细胞系的建立：通过在不含血清和细胞因子的合成培养基中培养人癌细胞系至少6个月进行选择后，从人癌细胞系如胃、结肠和卵巢细胞系建立了31个非血清依赖性生长细胞系。在无血清条件下建立的过程中，亲本细胞系显示出大致四种生长特性模式：1）在2周内，所有细胞均死亡。2)到4周时，细胞数量逐渐减少并消失。3)细胞只能在大量培养中生长。4)2.蛋白酶及其抑制剂在血清依赖性和非血清依赖性细胞系中的表达。通过酶谱、反向酶谱和Northern-blotting分析，建立了胃癌细胞系，并对其分泌的胰蛋白酶样酶进行了研究。经氨基酸序列测定，该酶为胰蛋白酶原-1。血清非依赖性细胞株中胰蛋白酶原-1 mRNA的表达量高于其亲本血清依赖性细胞株。3.血清和细胞因子对胰蛋白酶原-1 mRNA表达的调控。在评价细胞因子和生长因子对人胃癌细胞胰蛋白酶原-1基因表达的影响时，发现血清和炎性细胞因子对其他细胞系胰蛋白酶原-1基因的表达、胰蛋白酶原-1基因mRNA的表达具有调节作用。在筛选了多种人细胞系的胰蛋白酶原-1基因表达后，发现几种人癌细胞系如结肠癌和胃癌细胞系表达该基因的mRNA。然而，只有两个人胃癌细胞系受血清因子的调控。

1.Establishment of serum-independent human cell lines :After selection by culturing human cancer cell lines in the synthetic medium without serum nor cytokines for at least 6 months, 31 serum-independent growing cell lines were established from human cancer cell lines such as stomach, colon and ovarian cell lines. During establishment under serum-free conditions, parental cell lines showed roughly four patterns of growth properties : 1) within 2 weeks, all cells died simultaneiously. 2) by 4 weeks, cell number gradually decreased and disappeared. 3) cells could grow only in mass culture. 4) even from a single cell, it could gradually grow.2.Expression of proteinase and their inhibitors in serum-dependent and serum-independent cell lines. : By the analyzes using zymography, reversezymography and Northern-blotting, stomach cancer cell line established from malignant tumor secreted trypsin-like enzyme. The enzyme was identified to be trypsinogen-1 by its amino acid sequencing. And the enzyme was more expressed in serum-independent cell line than its parental serum-dependent cell line.3.Regulation of trypsinogen-1 mRNA expression by serum and cytokines. : When the effect of cytokines and growth factors on the expression fo trypsinogen-1 gene in human stomach cancer cells were evaluated, serum and inflammatory cytokines were revealed to regulate the expression of trypsinogen-1 gene.mRNA expression of trypsinogen-1 gene in other cell lines. : After screening various human cell lines on the expression of trypsinogen-1 gene, several human cancer cell lines such as colon and stomach cancer cell lines were revealed to express mRNA of the gene. However, only two human stomach cancer cell lines were regulated by serum factors.